The precise and reproducible development of the nervous system in vertebrates and invertebrates is accomplished by the directed growth of axons and dendrites (neurites) from neuronal cell bodies to their synaptic target cells. The extent and orientation of nerve growth is regulated and guided by a variety of molecules which are synthesized by neurons as well as non-neuronal cells. These regulators may be secreted or they may be immobilized on the surface of the cell which produces them. Binding of the regulator to a receptor on the neuronal cell surface causes a signal which regulates the intracellular molecules which control growth. Many types of molecules which regulate neuronal outgrowth are known. Some stimulate neurite growth (e.g., neurotrophic molecules, neurotransmitters, extracellular matrix molecules and cell adhesion molecules) while others function as inhibitors or negative regulators. The cell adhesion molecules (CAMs) are a large group of cell surface molecules which play an important role in regulating neuronal adhesion and neurite process extension. CAMs may be either Ca.sup.2+ -independent (e.g., N-CAM, contactin and L1/NgCAM) or Ca.sup.2+ -dependent (e.g., the cadherin protein family). Several of the Ca.sup.2+ -independent CAMs have been sequenced, and are structurally homologous to immunoglobulins. They are believed to mediate cell-cell adhesion by both homotypic and heterotypic binding mechanisms, and several are localized on the surfaces of axons during development. As several of the Ig homologues have been demonstrated to directly stimulate neurite outgrowth, this group of molecules is believed to play an important role in this process.
Of particular interest is the neural cell adhesion molecule, N-CAM, which serves as a ligand for homophilic adhesion between cells. N-CAM is generally recognized as a positive regulator of neuronal process outgrowth and is present in the membranes of developing neural cells and differentiated axons. Adult nerves, which normally express the adult A form of N-CAM, revert to expression of large amounts of the embryonic E form of this molecule upon injury. Antibodies to N-CAM have been shown to reduce the outgrowth of central and peripheral axons in vitro (B. R. Seckel. 1990. Muscle & Nerve 13: 785-800), and to disrupt reinnervation and functional recovery in transected sciatic nerves in vivo, presumably due to blockage of N-CAM binding (L. G. Remsen, et al. 1990. Exp. Neurol. 110:268-273).
When a nerve is severed, the regions of the neurites which are distal to the break become separated from the nerve cell body and degenerate. Death and degeneration of the neurites leaves only the empty nerve sheath and this, too, eventually degenerates. In addition there is some degeneration of the proximal stump. If degeneration does not result in the death of the nerve cell body it is possible for the nerves to regenerate by re-extension of the severed axons, especially if the regeneration occurs at a sufficient distance from the nerve cell body. The newly regenerating neurites are referred to as "nerve sprouts" and grow distally toward the sheath of the distal portion of the severed nerve. If the neurites successfully enter the sheath they will often grow down its length and function may be restored. Regrowth of the neurites is impeded or prevented by scar formation, which may be stimulated by trauma caused by suturing nerve ends in an effort to maintain alignment or by other manipulations involved in nerve repair. Nerve guide repair, which uses a conduit to bridge the gap between the proximal and distal ends of the severed nerve, has provided an alternative to nerve grafting and conventional repair techniques. Nerve guide repair is also referred to as "entubulation repair." Originally, entubulation repair employed an empty plastic tube applied to the damaged nerve ends to guide regrowth. With this technique there is less trauma, as only one epineural suture in each nerve stump is usually required to hold the nerve guide in place. Nerve guide repair may also prevent or reduce ingrowth of scar tissue which may interfere with the distal migration of the nerve sprout. The proximal stump, suspended in the nerve guide, may therefore begin its distal migration without obstruction by an imperfectly aligned degenerating distal stump, scar tissue, etc.
As certain cell adhesion molecules (CAMs, e.g., N-CAM, L1 Antigen, N-cadherin, GP135), neurotrophic factors (i.e., factors which promote survival and growth of neurons such as NGF and CNTF), and neurite-promoting factors (i.e., substrate bound glycoproteins which are usually components of the basal lamina such as laminin, collagen, entactin proteoglycan, fibrinogen and fibronectin) have been found to be involved in peripheral nerve regeneration, they have been used to modify nerve guides to improve nerve repair. Madison, et al. (1988. Brain Research 447:325-334) describe a nerve guide having a lumen filled with a collagen- or laminin-containing gel. U.S. Pat. No. 5,019,087 discloses a nerve guide having walls comprising Type I collagen or laminin. U.S. Pat. No. 5,011,486 discloses nerve guides comprising porous tubular membranes having a growth-enhancing active factor incorporated within the membrane. U.S. Pat. No. 4,759,764 discloses the use of basal lamina with the cellular material removed as a nerve guide. U.S. Pat. No. 4,955,892 discloses addition of a neural cell adhesion molecule (N-CAM) to a matrix in the lumen of the guide. While N-CAM enhances nerve growth, antibodies to N-CAM reportedly disrupted recovery of muscle function when nerve guides containing such antibodies were applied to transected sciatic nerves (L. G. Remsen, et al., supra). U.S. Pat. No. 5,026,381 discloses a nerve guide with multilayered, semipermeable walls comprising Type I collagen.
In the tubular nerve guides described in the prior art, the regenerating axons grow along the inside wall of the tube, along other axons, or along Schwann cells which have previously bridged the gap between the stumps. It is therefore likely that the axon will lose positional information and waste energy by growing in nonproductive directions as a large surface area is available for regrowth. This problem can be overcome by providing a nerve guide in the form of a fiber (i.e., a fiber nerve guide) which presents less surface area to the regenerating axon. The neurite therefore has less opportunity to lengthen its path to the target distal stump or sheath, resulting in more efficient regeneration. The fiber may be a surgical suture which is used to join a nerve bundle or fasicle of one stump of a severed nerve to the corresponding bundle of the other stump. Alternatively, the fiber nerve guide may be used in vitro to guide nerve cell outgrowth, in which case nerve cells may be placed on the fiber either as a mass of cells (e.g., a ganglion) or as dissociated cells useful for studies of individual nerve cells. A molecule which regulates neuronal process (nerve sprout) outgrowth may be applied to the surface of the fiber, i.e., layered thereon or covalently coupled to the fiber, to promote and direct this process.